GUANIDINOACETATE AMIDINOHYDROLASE FROM A FLAVOBACTERIUM PURIFICATION, CRYSTALLIZATION AND PROPERTIES

Guanidinoacetate amidinohydrolase (glycocyaminase, EC 3.5.3.2) was purified 148-fold and crystallized from Flavobacterium sp. GE-1 grown in a medium supplemented with guani- dinoacetate. The molecular weight of the enzyme was estimated to be about 281,000 daltons by a gel electrophoretic procedure. SDS-polyacrylamide electrophoresis showed a single component of subunit with an estimated molecular weight of 70,000 daltons, suggesting that the enzyme is composed of four subunits identical in molecular weight. These values are significantly distinct from those of an analogous enzyme from a Pseudomonas reported by us previously. The native enzyme was active without added divalent metals. Incubation of the enzyme with 1,10-phenanthroline at an elevated temperature (50°C) resulted in almost complete inactivation. The activity of the inactivated enzyme was restored by incubation with Zn2+ or Co2+at 50°C; the former cation was more effective. The enzyme was optimally active at pH 8.0 - 8.5. The apparent Km value for guanidinoacetate was 15 mM. 3-Guanidinopropionate and 4-guanidinobutyrate were hydrolyzed 19% and 9%, respectively, as fast as guanidinoacetate. PCMB was a potent inhibitor of the enzyme. The properties of the Flavobacterium enzyme were compared with those of the analogous Pseudomonas enzyme. © 1979, by the Agricultural Chemical Society of Japan.